The main objective of this project is to study the mechanism of carcinogenesis using quantitative two-dimensional gel electrophoresis. Research is, at present, focused on the following areas: (1) continued development and improvements on the computer system (dubbed ELSIE III) currently used to automatically analyze two-dimensional gels, and (2) use of ELSIE III to analyze experiments requiring computerized analysis of two-dimensional gel electrophoretograms. In the past year emphasis has been placed on developing software tools to aid the investigator in identifying spots of interest in an experiment. Statistical tests have been incorporated into various program designed to search through sets of matched spots. These tests can be used to search for spots that may vary quantitatively or qualitatively over the course of an experiment. Once such spots are flagged, interactive computer graphics can be used to examine the spots and confirm their relevance to the experiment. The system has been used in several studies in this and other laboratories. One study involves the detailed quantitative evaluation of modulation in the rates of protein synthesis in the rat hepatoma cell line, H4-II-E (Reuber cells). Several single-cell-derived cultures of H4-II-E were prepared and grown under identical conditions. Cells were labeled as soon as possible after the cultures were established and two-dimensional gels were run on a total cell lysate. Quantitative analysis showed that 10% or more of the proteins demonstrate statistically significant modulation in their rates of synthesis when compared from culture to culture. These differences were generally on the order of 50%, although changes of as much as fourfold were detected. We conclude tht the rates of synthesis of many polypeptides can vary significantly in a culture as a result of very subtle (and apparently uncontrollable) variations in the environment of cells and that the ELSIE III system is capable of detecting these changes.